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Introduction

While most in vitro assays are designed to study a particular step in the angiogenesis process, the aortic ring assay is unique in that it recapitulates all of the key steps in the process (matrix degradation, migration, proliferation, reorganization).

Assay Description

Originally developed by Nicosia et al ¹, the rat thoracic aorta is excised, the connective tissue is removed, and small ring-like segments are cut and embedded into a three-dimensional matrix composed of fibrin or collagen and maintained in a chemically-defined medium. New blood vessel growth, triggered by the injury of the dissection procedure and mediated by growth factors produced from the explant, can be observed several days after culture initiation. Test substances added to the culture can be analyzed for their ability to positively or negatively affect vessel growth. Image analysis software is normally used to measure the length and the abundance of these sprouting vessels.

This in vitro assay has several advantages:1). all of the key steps of the angiogenesis process are represented;2) multiple cell types are represented; and3) the endothelial cells within the aortic explant have not been pre-selected by passaging and therefore are not in a proliferative state, which is closer to the real-life environment in which angiogenesis occurs.

Disadvantages include the variability that is possible within an experiment, due to the incomplete removal of the outer connective tissue surrounding the aorta, which can influence microvessel outgrowth. Variability in angiogenic responses is also possible among multiple rat aortas. Another difficulty a user typically faces is quantification of microvessel outgrowth, which can occur in three dimensions. Lastly, while the use of the aorta is a practical choice, it is not ideal because in vivo angiogenesis typically occurs in microvessels but not major vessels like the aorta.